Hypoglycemic agent

ABSTRACT

A zinc(II) complex which is lowly toxic, has high insulin-like activity, and is effectively usable as a hypoglycemic agent for the prevention or treatment of diabetes; a hypoglycemic agent containing the complex; a medicinal preparation which contains the complex and is useful as a preventive/remedy for diabetes; and a food containing the complex, such as a health food or supplementary health food. The hypoglycemic agent contains an organic zinc(II) complex having as a ligand a compound selected among aminoalkylpyridines, bis-optically active amino acids, bisaminoalkylcarboxylic acids, oligopeptides, oligopseudopeptides, di-substituted aminocarboxylic acids, α- and β-hydroxycarboxylic acids, vitamins, glutamine derivatives, etc.

TECHNICAL FIELD

The present invention relates to a novel hypoglycemic agent which hasinsulin-like activity and is useful as a preventive/remedy for diabetes.Precisely, the invention relates to a novel hypoglycemic agent thatcontains a zinc(II) complex having a specific compound as a ligand andhaving insulin-like activity.

BACKGROUND ART

At present, treatment of type I (insulin-dependent) diabetes inevitablydepends on subcutaneous insulin injection, and it is desired to developan oral remedy for it in place of insulin. Some remedies have beendeveloped for type II (insulin-independent) diabetes derived fromstress, obesity, lack of exercise, ageing or the like, and have beentried in clinical treatment. However, none of them is a panacea and someoften cause a problem of side effects. One of such remedies is vanadylsulfate, and it has already been tried in clinical treatment in USA etc.In addition, vanadyl sulfate and bispicolinic acid/vanadyl complex arecommercially available in USA as supplementary health foods.

On the other hand, it has been known that zinc(II) ion, which is knownto be less toxic than vanadium, has insulin-like activity since around1980 (L. Coulston and P. Dandona, “Insulin-like effects of Zn²⁺ onadipocytes”, Diabetes, 29, 665-7 (1980); J. M. May and C. S. Contoreggi,“The mechanism of the insulin-like effects of ionic zinc”, J. Biol.Chem., 257, 4362-8 (1982), and A. Shisheva, D. Gefel and Y. Shechter,“Insulin-like effects of zinc ion in vitro and in vivo” (Zn²⁺ is thefirst agent other than vanadate that on oral administration is able torestore tissue ability to metabolism glucose), Diabetes, 41, 982-8(1992)). Since vanadyl sulfate and zinc(II) ion (zinc sulfate and zincchloride) are inorganic salts, they are hardly permeable throughbio-membranes and are therefore hardly taken into living bodies. Toovercome the problems, zinc(II) complexes, which are less toxic thanvanadium and are favorably stable and fat-soluble and have insulin-likeactivity, may be more effective than vanadyl complexes, and developingthem is desired.

On the other hand, zinc that is contained in brewer's yeast and seaweedextracts is commercially available in Japan as supplementary healthfoods.

Regarding zinc complexes, the present inventors have already filed apatent application that has been already published, InternationalPublication WO01/39769A1 (international publication date: Jun. 7, 2001)“Hypoglycemic Agent Comprising Zinc(II) Organic Complex”; and somereports have been announced (for example, Y. Yoshikawa, E. Ueda, Y.Suzuki, N. Yanagihara, H. Sakurai and Y. Kojima, “New InsulinomimeticZinc(II) Complexes of α-Amino Acids and their Dervatives with Zn(N₂O₂)Coordination Mode”, Chem. Pharm. Bull., 49, 652-654 (2001); Yoshikawa,Ueda, Sakurai and Kojima, “Development and Study of Zinc(II) Complexeswith Hypoglycemic Activity”, Biomed Res Trace Elements, 12, 104-109(2001)). However, it is desired to further develop more effectivehypoglycemic agents comprising a complex that is less toxic and hashigher activity and to develop foods such as health foods andsupplementary health foods having the effect.

DISCLOSURE OF THE INVENTION

The present invention has been made in consideration of theabove-mentioned current situation, and it aims to provide a zinc(II)complex that is less toxic, has high insulin-like activity and iseffectively usable as a hypoglycemic agent for the prevention ortreatment of diabetes, to provide a hypoglycemic agent that contains thesaid complex, to provide a medicinal preparation that contains the saidcomplex and is useful as a preventive/remedy for diabetes, and toprovide foods such as health foods and supplementary health food thatcontain the said complex.

The invention relates to a hypoglycemic agent that contains a zinc(II)organic complex having, as a ligand, any compound selected from thefollowing (1) to (12):

(1) compounds of a general formula (1) (when they are optically-activecompounds, they contain both (R)-form and (S)-form thereof):

(wherein R, R′, R¹ and R² each independently represents a hydrogen atom,an alkyl group, an aryl group, or an aralkyl group; n indicates aninteger of from 1 to 3);

(2) compounds of a general formula (2):

(wherein X¹ and X² each independently represents an alkoxy group, anamino group, a mono-lower alkylamino group, a di-lower alkylamino group,or a hydroxyl group; R and R′ each independently represents a hydrogenatom, an alkyl group, an aryl group, or an aralkyl group; R³ and R⁴ eachindependently represents an alkyl group, a substituted alkyl group, or aheterocyclic group; R and R³, and/or R′ and R⁴ may be taken together toform an alkylene group; C* represents an asymmetric carbon (either(R)-form or (S)-form); and X represents an alkylene group);

(3) compounds of a general formula (3):

(wherein X¹ and X² each independently represents an alkoxy group, anamino group, a mono-lower alkylamino group, a di-lower alkylamino group,or a hydroxyl group; R and R′ each independently represents a hydrogenatom, an alkyl group, an aryl group, or an aralkyl group; X representsan alkylene group; n indicates an integer of from 1 to 3);

(4) compounds of a general formula (4) (when they are optically-activecompounds, they contain both (R)-form and (S)-form thereof):

(wherein X¹ represents an alkoxy group, an amino group, a mono-loweralkylamino group, a di-lower alkylamino group, or a hydroxyl group; R,R′ and R″ each independently represents a hydrogen atom, an alkyl group,an aryl group, or an aralkyl group; R^(3′) and Ro³ each independentlyrepresents a hydrogen atom, an alkyl group, a substituted alkyl group,or a heterocyclic group; R and R^(3′), and/or R″ and Ro³ may be takentogether to form an alkylene group; n indicates an integer of from 1 to3);

(5) compounds of a general formula (5) (when they are optically-activecompounds, they contain both (R)-form and (S)-form thereof):

(wherein X¹ represents an alkoxy group, an amino group, a mono-loweralkylamino group, a di-lower alkylamino group, or a hydroxyl group; Rand R′ each independently represents a hydrogen atom, an alkyl group, anaryl group, or an aralkyl group; R^(3′), R^(4′) and R⁵ eachindependently represents a hydrogen atom, an alkyl group, a substitutedalkyl group, or a heterocyclic group; R and R^(3′) may be taken togetherto form an alkylene group; n indicates an integer of from 1 to 3);

(6) compounds of a general formula (6) (when they are optically-activecompounds, they contain both (R)-form and (S)-form thereof):

[wherein X¹ represents an alkoxy group, an amino group, a mono-loweralkylamino group, a di-lower alkylamino group, or a hydroxyl group;R^(3′) represents a hydrogen atom, an alkyl group, a substituted alkylgroup, or a heterocyclic group; R⁶ and R⁷ each independently representsan alkyl group, an aralkyl group, or a group of the following formula(6′):

(wherein R⁸ and R⁹ each independently represents a hydrogen atom, analkyl group, a nitro group, or a halogen atom); R⁶ and R^(3′) may betaken together to form an alkylene group];

(7) compounds of a general formula (7):

[wherein R¹⁰ and R¹¹ each independently represents a hydrogen atom, analkyl group, a hydroxyalkyl group, —(CH₂)_(m)NRR′, —(CH₂)_(m)N⁺RR′R″,—(CH₂)_(m)SR, —(CH₂)_(m)S⁺RR′, —(CH₂)_(m)COOR or —(CH₂)_(m)CONRR′ (wherem indicates an integer of from 0 to 4; and R, R′ and R″ eachindependently represents a hydrogen atom, an alkyl group, an aryl group,or an aralkyl group)];

(8) compounds of a general formula (8):

(9) compounds of a general formula (9):

[wherein R¹⁰ and R¹¹ each independently represents a hydrogen atom, analkyl group, a hydroxyalkyl group, —(CH₂)_(m)NRR′, —(CH₂)_(m)N⁺RR′R″,—(CH₂)_(m)SR, —(CH₂)_(m)S⁺RR′, —(CH₂)_(m)COOR or —(CH₂)_(m)CONRR′ (wherem indicates an integer of from 0 to 4; and R, R′ and R″ eachindependently represents a hydrogen atom, an alkyl group, an aryl group,or an aralkyl group)];

(10) compounds of a general formula (10):

(11) compounds of a general formula (11):

(wherein X_(o) ⁻ represents Cl⁻, Br⁻ or I⁻);

(12) compounds of a general formula (12):

(wherein R¹² and R¹³ each independently represents an alkyl group, anaryl group, or an aralkyl group).

The invention also relates to an oral formulation comprising thezinc(II) organic complex that contains, as a ligand, any compoundselected from the above-mentioned general formulae (1) to (12).

Further, the invention relates to a pharmaceutical compositioncomprising, as an active ingredient, the zinc(II) organic complex thatcontains, as a ligand, any compound selected from the above-mentionedgeneral formulae (1) to (12).

Still further, the invention relates to zinc(II) organic complexeshaving, as a ligand, a compound of the above-mentioned general formula(2), (3), (4), (5), (6), (9), (10), (11) or (12).

The invention also relates to a food that contains a zinc(II) organiccomplex having, as a ligand, any of L-lactic acid, quinic acid,L-carnitine (vitamin B_(T)), vitamin C, vitamin U or L-theanine.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the effect of zinc(II) organic complexes of the inventionon release of fatty acids from fat cells. In FIG. 1, 1 indicates theresult of a blank, 2 indicates the result of a control, 3 to 5 indicatethe results of positive controls, and 6 to 20 indicate the results ofthe compounds of the invention.

FIG. 2 shows the effect of zinc(II) organic complexes of the inventionon release of fatty acids from fat cells. In FIG. 2, 1 indicates theresult of a blank, 2 indicates the result of a control, 3 to 5 indicatethe results of positive controls, and 6 to 20 indicate the results ofthe compounds of the invention.

FIG. 3 shows the effect of zinc(II) organic complexes of the inventionon release of fatty acids from fat cells. In FIG. 3, 1 indicates theresult of a blank, 2 indicates the result of a control, 3 to 5 indicatethe results of positive controls, and 21 to 38 indicate the results ofthe compounds of the invention.

FIG. 4 shows a full chart of the IR absorption spectrum (IR) ofbis(2-aminomethylpyridine)/zinc(II) complex [Zn(2-AM-py)₂(H₂O)Cl₂] ofthe invention.

FIG. 5 shows a full chart of the IR absorption spectrum (IR) ofL-carnitine/zinc(II) complex [Zn(Car)₂Cl₂] of the invention.

FIG. 6 shows a full chart of the IR absorption spectrum (IR) ofbis(L-ascorbic acid)/zinc(II) complex [Zn(Vit-C)₂] of the invention.

FIG. 7 shows a full chart of the IR absorption spectrum (IR) ofbis(vitamin U)/zinc(II) complex [Zn(Vit-U)Cl₂] of the invention.

FIG. 8 shows a full chart of the IR absorption spectrum (IR) ofbis(L-theanine)/zinc(II) complex [Zn(Tea)₂] of the invention.

FIG. 9 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of Zn(2-AM-py)₂Cl₂ for 14 days (-o-) andwith no administration thereof (control) (-•-).

FIG. 10 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of Zn(Lac)₂ for 14 days (-o-) and with noadministration thereof (control) (-•-).

FIG. 11 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of Zn(Qui)₂ for 14 days (□) and with noadministration thereof (control) (▪).

FIG. 12 shows the blood glucose level (BGL) change in KK-A^(y) mice withoral administration of Zn(Car)₂Cl₂ for 14 days (-o-) and in those withoral administration of L-carnitine for 14 days (-♦-).

FIG. 13 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of a solution (about pH 7) prepared bymixing zinc sulfate and vitamin U in a molar ratio of 1:2 (that is, asolution of Zn(Vit-U)Cl₂)) for 14 days (-o-) and with no administrationthereof (control) (-•-).

FIG. 14 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of Zn(2-AM-py)₂Cl₂ for 14 days (-o-) andwith no administration thereof (control) (-•-).

FIG. 15 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of Zn(Lac)₂ for 14 days (-o-) and with noadministration thereof (control) (-•-).

FIG. 16 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of Zn(Qui)₂ for 14 days (-o-) and with noadministration thereof (control) (-•-).

FIG. 17 shows the body weight change of KK-A^(y) mice with oraladministration of Zn(Car)₂Cl₂ for 14 days (-o-) and of those with oraladministration of L-carnitine for 14 days (-♦-).

FIG. 18 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of a solution (about pH 7) prepared bymixing zinc sulfate and vitamin U in a molar ratio of 1:2 (that is, asolution of Zn(Vit-U)Cl₂)) for 14 days (-o-) and with no administrationthereof (control) (-•-).

FIG. 19 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with intraperitoneal administration ofZn(2-AM-py)₂Cl₂ for 14 days (-o-) and with no administration thereof(control) (-•-). The glucose tolerance test was effected after the 14days administration.

FIG. 20 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with oral administration of Zn(Car)₂Cl₂ for 14days (-o-), with oral administration of L-carnitine for 14 days (-♦-),and with administration of acacia solution alone for 14 days (control)(-•-). The glucose tolerance test was effected after the 14 daysadministration.

FIG. 21 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with intraperitoneal administration of Zn(Qui)₂for 14 days (-o-) and with no administration thereof (control) (-•-).The glucose tolerance test was effected after the 14 daysadministration.

FIG. 22 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with intraperitoneal administration of a solution(about pH 7) prepared by mixing zinc sulfate and vitamin U in a molarratio of 1:2 (that is, a solution of Zn(Vit-U)Cl₂)) for 14 days (-o-)and with no administration thereof (control) (-•-). The glucosetolerance test was effected after the 14 days administration.

BEST MODES OF CARRYING OUT THE INVENTION

The ligand for the zinc(II) organic complexes of the invention includesthe compounds of the above-mentioned general formulae (1) to (12).

The compounds of formula (1) may be referred to, for example, asaminoalkylpyridines.

The compounds of formula (2) may be referred to, for example, asbis-optical active amino acids.

The compounds of formula (3) may be referred to, for example, asbisaminoalkylcarboxylic acids.

The compounds of formula (4) may be referred to, for example, asoligopeptides.

The compounds of formula (5) may be referred to, for example, asoligo-pseudopeptides.

The compounds of formula (6) may be referred to, for example, asdi-substituted aminocarboxylic acids.

The compounds of formula (7) and those of formula (8) may be referredto, for example, as α-oxycarboxylic acids.

The compounds of formula (9) may be referred to, for example, asβ-oxycarboxylic acids.

Of the compounds of formula (9), L-carnitine is vitamin B_(T); theL-form compound of formula (10) is vitamin C; and the L-form compound offormula (11) is vitamin U.

R, R′, R¹ and R² in formula (1), R and R′ in formula (2), R and R′ informula (3), R, R′ and R″ in formula (4), and R and R′ in formula (5)each independently represents a hydrogen atom, an alkyl group, an arylgroup or an aralkyl group. For these, the alkyl group is, for example, alinear or branched lower alkyl group having from 1 to 6, preferably from1 to 4 carbon atoms. More concretely, it includes, for example, a methylgroup, an ethyl group, a propyl group, an isopropyl group, a butylgroup, an isobutyl group, a secondary butyl group, and a tertiary butylgroup. The aryl group includes, for example, a phenyl group, a tolylgroup, a xylyl group, and a naphthyl group. The aralkyl group includes,for example, a benzyl group, a phenethyl group, a naphthylmethyl group,and a naphthylethyl group.

R³ and R⁴ in formula (2) each independently represents an alkyl group, asubstituted alkyl group, or a heterocyclic group; and R^(3′) and Ro³ informula (4), R^(3′), R^(4′) and R⁵ in formula (5), and R^(3′) in formula(6) each independently represents a hydrogen atom, an alkyl group, asubstituted alkyl group, or a heterocyclic group. For these, the alkylgroup is, for example, a linear or branched lower alkyl group havingfrom 1 to 6, preferably from 1 to 4 carbon atoms. More concretely, forexample, it is preferably a methyl group, an isopropyl group, anisobutyl group, or a secondary butyl group. The substituted alkyl groupincludes, for example, those derived from a lower alkyl group such as amethyl, ethyl or propyl group by substituting the hydrogen atom thereofwith any of a hydroxyl group, a carboxyl group, an amino group, an amidogroup, a thiol group, a methylthio group, a phenyl group, ahydroxyphenyl group, or —NHC(NH₂)═NH. Concretely, for example, itincludes —CH₂OH, —CH(CH₃)OH, —(CH₂)₃NHC(NH₂)═NH, —(CH₂)₄NH₂,—(CH₂)₂S(CH₃), —CH₂COOH, —CH₂CONH₂, —(CH₂)₂COOH, —(CH₂)₂CONH₂, —CH₂SH, abenzyl group, and a 4-hydroxyphenylmethyl group. The heterocyclic groupis, for example, a saturated or unsaturated, monocyclic, polycyclic orcondensed cyclic group having at least one of nitrogen, oxygen andsulfur atoms in the ring, of which one ring is 5- to 20-membered,preferably 5- to 10-membered, more preferably 5- to 7-membered and whichmay be condensed with a carbon cyclic group such as a cycloalkyl group,a cycloalkenyl group or an aryl group. Preferred examples of the groupare an imidazolyl group and an indolyl group.

The alkylene group to be formed by R and R³, R and R^(3′), R′ and R⁴, R″and Ro³, and R⁶ and R^(3′) includes, for example, an ethylene group, apropylene group, and a butylene group.

X¹ and X² in formula (2), X¹ and X² in formula (3), X¹ in formula (4),X¹ in formula (5), and X¹ in formula (6) each independently representsan alkoxy group, an amino group, a mono-lower alkylamino group, adi-lower alkylamino group, or a hydroxyl group. The alkoxy group forthese is, for example, a linear or branched lower alkoxy group havingfrom 1 to 6, preferably from 1 to 4 carbon atoms. More concretely, forexample, it includes a methoxy group, an ethoxy group, a propoxy group,an isopropoxy group, a butoxy group, an isobutoxy group, a secondarybutoxy group, and a tertiary butoxy group. The mono-lower alkylaminogroup is a substituted mono-lower alkylamino group with, for example, alinear or branched lower alkyl group having from 1 to 6, preferably from1 to 4 carbon atoms. More concretely, for example, it includes amethylamino group, an ethylamino group, a propylamino group, anisopropylamino group, a butylamino group, and a tertiary butylaminogroup. The di-lower alkylamino group is a substituted di-loweralkylamino group with, for example, a linear or branched lower alkylgroup having from 1 to 6, preferably from 1 to 4 carbon atoms. Moreconcretely, for example, it includes a dimethylamino group, adiethylamino group, a dipropylamino group, a diisopropylamino group, adibutylamino group, and a di-tertiary butylamino group.

R⁶ and R⁷ in formula (6) each independently represents an alkyl group,an aralkyl group, or a group of the following formula (6′):

(wherein R⁸ and R⁹ each independently represents a hydrogen atom, analkyl group, a nitro group, or a halogen atom). The alkyl group forthese is, for example, a linear or branched lower alkyl group havingfrom 1 to 6, preferably from 1 to 4 carbon atoms. More concretely, itincludes, for example, a methyl group, an ethyl group, a propyl group,an isopropyl group, a butyl group, an isobutyl group, a secondary butylgroup, and a tertiary butyl group. The aralkyl group includes, forexample, a benzyl group, a phenethyl group, a naphthylmethyl group, anda naphthylethyl group. The halogen atom includes, for example, chlorine,bromine, iodine and fluorine atoms.

X in formulae (2) and (3) represents an alkylene group. The alkylenegroup is, for example, a lower alkylene group having from 1 to 6,preferably from 1 to 4 carbon atoms. More concretely, for example, itincludes an ethylene group, a trimethylene group, a methylethylenegroup, a propylene group, a tetramethylene group, and a1,2-dimethylethylene group.

R¹⁰ and R¹¹ in formulae (7) and (9) each independently represents ahydrogen atom, an alkyl group, a hydroxyalkyl group, —(CH₂)_(m)NRR′,—(CH₂)_(m)N⁺RR′R″, —(CH₂)_(m)SR, —(CH₂)_(m)S⁺RR′, —(CH₂)_(m)COOR or—(CH₂)_(m)CONRR′ (where m indicates an integer of from 0 to 4; and R, R′and R″ each independently represents a hydrogen atom, an alkyl group, anaryl group, or an aralkyl group). For these, the alkyl group is, forexample, a linear or branched lower alkyl group having from 1 to 6,preferably from 1 to 4 carbon atoms. More concretely, it includes, forexample, a methyl group, an ethyl group, a propyl group, an isopropylgroup, a butyl group, an isobutyl group, a secondary butyl group, and atertiary butyl group. The hydroxyalkyl group is, for example, theabove-mentioned alkyl group of which one hydrogen atom is substitutedwith a hydroxyl group. More concretely, for example, it includes ahydroxymethyl group, an α-hydroxyethyl group, a β-hydroxyethyl group, ahydroxypropyl group, and a hydroxybutyl group. In the other groups, R,R′ and R″ each independently represents a hydrogen atom, an alkyl group,an aryl group, or an aralkyl group. For the alkyl group, the aryl groupand the aralkyl group, referred to are the same mentioned hereinabovefor R, R′, R¹ and R² in formula (1).

R¹² and R¹³ in formula (12) each independently represents an alkylgroup, an aryl group or an aralkyl group. For the alkyl group, the arylgroup and the aralkyl group, referred to are the same mentionedhereinabove for R, R′, R¹ and R² in formula (1).

The zinc(II) organic complexes of the invention may be produced by knownmethods (for example, as in U.S. Pat. No. 5,219,847) or according tosuch known methods. For example, a solution of a zinc salt is added to asolution of the intended ligand to form a zinc(II) organic complex, andwhich is then isolated. For the solvent, water is generally preferred,but an organic solvent or a mixed solvent may also be used. The zincsalt solution is preferably an aqueous solution of an inorganic zincsuch as zinc sulfate, zinc nitrate or zinc chloride. Preferably, the pHof the reaction solution is controlled, if desired. For thepH-regulating agent, usable is an aqueous basic solution of, forexample, sodium hydroxide, lithium hydroxide, potassium hydroxide orbarium hydroxide.

More concretely, referred to are Examples described hereinunder.

Of the zinc(II) organic complexes of the invention, those having, as aligand, a compound of formula (2), (3), (4), (5), (6), (9), (10), (11)or (12) are all novel compounds.

As is obvious from Test Examples to be described hereinunder, thezinc(II) organic complexes of the invention have insulin-like activityor hypoglycemic activity, and are useful as preventives/remedies fordiabetes and hypertension.

Accordingly, the invention provides a pharmaceutical composition thatcomprises the above-mentioned zinc(II) organic complex of the inventionand a pharmaceutically-acceptable carrier.

The pharmaceutical composition may be in the form of a medicinalpreparation that comprises the zinc(II) organic complex of the inventionserving as an active ingredient, along with apharmaceutically-acceptable carrier such as an organic or inorganic,solid or liquid vehicle suitable for oral administration, parenteraladministration or local administration. The medicinal preparation may bein any form of capsules, tablets, pills, granules, powders, inhalants,suppositories, solutions, lotions, suspensions, emulsions, ointments andgels. If desired, the preparation may contain an auxiliary aid, astabilizer, a wetting agent, a emulsifier, a buffer and any otherordinary additives.

The invention provides an oral formulation that contains the zinc(II)organic complex. The oral formulation of the invention contains thezinc(II) organic complex having insulin-like activity or hypoglycemicactivity. Accordingly, the invention provides an oral remedy/preventivefor diabetes. The invention also provides use of the zinc(II) organiccomplex for producing such an oral preventive/remedy for diabetes, andprovides a method for preventing and treating diabetes by orallyadministering an effective dose of the oral preventive/remedy fordiabetes.

The effective dose of the zinc(II) organic complex of the invention forprevention/treatment of the disease varies depending on the age and thecondition of the cases to which it is administered. In general, however,a mean dose of the zinc(II) organic complex of the invention may fallbetween about 0.1 mg/patient and about 1000 mg/patient, and it may beadministered once or a few times a day.

FIG. 1, FIG. 2 and FIG. 3 show test results that indicate the capabilityof the zinc(II) organic complex of the invention to inhibit the releaseof fatty acids from the fat cells in rats.

FIG. 1, FIG. 2 and FIG. 3 each shows the free fatty acid inhibitingeffect of the zinc(II) organic complex of the invention added toepinephrine-stimulated rat fat cells.

In FIG. 1, 1 indicates a blank, 2 indicates a control, 3 to 5 indicatepositive controls with oxovanadium sulfate (VOSO₄), and 6 to 20 indicatethe compounds of the invention.

6 to 8 are with N,N′-ethylene-bis-β-alanine/zinc(II) complex[Zn(βAeAβ)(H₂O)₂]; 9 to 11 are withN,N′-trimethylene-bis-L-valine/zinc(II) complex [Zn(VtV)(H₂O)₂]; 12 to14 are with N,N-dipyridylmethyl-L-valine/zinc(II) complex[Zn(^(pm2)V)(ClO₄)]; 15 to 17 are withN,N-dipyridylmethyl-D-valine/zinc(II) complex [Zn(^(pm2)V_(R))(ClO₄)];18 to 20 are with N-6-methylpyridylmethyl-L-aspartic acid/zinc(II)complex [Zn(^(6Me-pm)D)].

The concentration of the compound used is 10⁻⁴ M, 5×10⁻⁴ M, and 10⁻³ M,respectively.

In FIG. 2, 1 indicates a blank, 2 indicates a control, 3 to 5 indicatepositive controls with oxovanadium sulfate (VOSO₄), and 6 to 20 indicatethe compounds of the invention.

6 to 8 are with bis(2-aminomethylpyridine)/zinc(II) complex[Zn(2-AM-py)₂(H₂O)Cl₂); 9 to 11 are withbis-(R)-2-(1-aminoethyl)pyridine (=α-pyridylethylamine)/zinc(II) complex[Zn((R)-2-(1-AE)-py)₂(H₂O)Cl₂); 12 to 14 are withbis-(S)-2-(1-aminoethyl)pyridine/zinc(II) complex[Zn((S)-2-(1-AE)-py)₂(H₂O)Cl₂); 15 to 17 are withL-aspartyl-L-phenylalanine methyl ester (aspartame)/zinc(II) complex[Zn(AspF—OMe)(H₂O)₂]; 18 to 20 are withbis(glycyl-N,N′-ethylene-L-alanyl-L-alanine ethyl ester)/zinc(II)complex [Zn(Gly-eAA-OEt)₂Cl₂].

The concentration of the compound used is 10⁻⁴ M, 5×10⁻⁴ M, and 10⁻³ M,respectively.

Further, in FIG. 3, 1 indicates a blank, 2 indicates a control, 3 to 5indicate positive controls with oxovanadium sulfate (VOSO₄), and 21 to38 indicate the compounds of the invention.

21 to 23 are with bis(L-lactic acid)/zinc(II) complex [Zn(Lac)₂]; 24 to26 are with bis(quinic acid)/zinc(II) complex [Zn(Qui)₂]; 27 to 29 arewith bis(L-carnitine)/zinc(II) complex [Zn(Car)₂Cl₂]; 30 to 32 are withbis(L-ascorbic acid)/zinc(II) complex [Zn(Vit-C)₂]; 33 to 35 are withbis(vitamin U)/zinc(II) complex [Zn(Vit-U)Cl₂]; and 36 to 38 are withbis(L-theanine)/zinc(II) complex [Zn(Tea)₂].

The concentration of the compound used is 10⁻⁴ M, 5×10⁻⁴ M, and 10⁻³ M,respectively.

The “blank” 1 in FIG. 1, FIG. 2 and FIG. 3 shows the value of free fattyacids (FFA) in spontaneous release from cells; and the “control” 2therein shows the value thereof released through epinephrinestimulation. 3 to 5 with oxovanadium sulfate “VOSO₄” in FIG. 1, FIG. 2and FIG. 3 are comparative examples.

Table 1 below shows IC₅₀ (mM), the concentration of the test compound,zinc(II) complex of the invention that inhibits 50% fatty acid releasecalculated based on the results of the tests. In Table 1, IC₅₀ (mM) of“VOSO₄” is 1.00 mM, and the value of the other compounds is a relativevalue to it.

TABLE 1 Complex IC₅₀ (mM) Zn(2-AM-py)₂Cl₂ 0.85 Zn((R)-2-AM-py)₂Cl₂ 0.40Zn((S)-2-AM-py)₂Cl₂ 0.39 Zn(βAeAβ) 0.82 Zn(VtV) 0.92 Zn(AspF-OMe)₂ 1.11Zn(Gly-eAA-OEt)₂Cl₂ 0.85 Zn(^(pm2)V)ClO₄ 0.79 Zn(^(pm2)V_(R))ClO₄ 0.92Zn(^(6Me-pm)D) 0.86 Zn(Lac)₂ 0.81 Zn(Qui)₂ 0.98 Zn(Car)₂Cl₂ 0.80Zn(Vit-C)₂ 0.80 Zn(Vit-U)Cl₂ 0.84 Zn(Tea)₂ 1.23 VOSO₄ 1.00

The data confirms that, as compared with VOSO₄, the zinc(II) organiccomplex of the invention significantly inhibits fatty acids from beingreleased from rat fat cells, indicating that the complex is favorablefor preventives/remedies for diabetes and hypertension.

In addition, the zinc(II) organic complex of the invention is usable infoods such as health foods, supplementary health foods, nutrient foodsand supplementary nutrient foods that have hypoglycemic activity.

In particular, the zinc(II) organic complex of the invention that has anatural ligand such as L-lactic acid, quinic acid, L-carnitine (vitaminB_(T)), vitamin C, vitamin U or L-theanine is especially favorable forfoods such as health foods, supplementary health foods, nutrient foodsand supplementary nutrient foods that have hypoglycemic activity.

The foods of the invention may contain any other foods, food additives,vitamins and/or minerals.

These other foods, food additives, vitamins and minerals may be any onesthat are generally used and may be used in future in the field ofmedicines and foods. Naturally, however, since the foods of theinvention are health foods, supplementary health foods, nutrient foods,supplementary nutrient foods and the like that have hypoglycemicactivity, the additional foods, food additives, vitamins and minerals tobe added thereto must not interfere with the effect of the foods of theinvention.

Regarding the shape thereof, the foods of the invention may be in anyform of powders, granules, tablets, capsules, liquids, gels and anyothers.

Not limited to those having the shape as above, foods and drinks thatare produced by adding the zinc(II) organic complex of the invention toalready-existing foods (foods and drinks) are all within the scope ofthe foods of the invention.

Examples of the foods and drinks are drinks such as refreshing drinks,nutrient drinks, fruit drinks, lactic acid drinks (includingconcentrated stocks and/or controlled powders to give these drinks);frozen confectionery such as ice creams, sherbets; processed cerealssuch as buckwheat noodles, wheat noodles, baked goods, rice cakes, doughfor jiaozi (Chinese dumpling with minced pork and vegetable stuffing);confectionery such as caramels, candies, chocolates, snacks, biscuits,cookies, crackers, jellies, jams; processed marine and stock farmproducts such as steamed fish pastes, pounded fish cakes, hams,sausages; milk products such as processed milks, cheeses, butters; oilsand fats, and processed oils and fats such as margarines, lards,mayonnaises; seasonings such as soy sauces, sauces, soybean pastes,juices pressed from bitter oranges, tangleweed extracts, soup stocks;various everyday dishes; pickles; and other various types ofsupplementary nutrient and health foods. Needless-to-say, these are notlimitative.

The foods of the invention, which comprise an organic compound capableof forming a complex with zinc and a zinc source, are much expected forhealth (supplementary) foods, more concretely, for example, supplements(specific health foods, so-called “tokuho”) that normalize the bloodglucose level of diabetics and others and are effective for preventionand treatment of glucose tolerance disorders, diabetes (e.g., type IIdiabetes), insulin-resistant syndromes (e.g., insulin receptordisorders), polycystic ovary syndromes, hyperlipemia, atherosclerosis,cardiovascular disorders (e.g., stenocardia, cardiac insufficiency),hyperglycemia, hypertension, stenocardia, pulmonary hypertension,congestive cardiac insufficiency, diabetic complications (e.g., diabeticgangrene, diabetic arthropathy, diabetic glomerulosclerosis, diabeticskin disorders, diabetic neuropathy, diabetic cataract, diabeticretinopathy), skin disorders, taste disorders, etc.

In addition, the foods of the invention are much expected for nutrient(supplementary) foods having the ability of insulin activation and bloodglucose level normalization.

EXAMPLES

The invention is described in more detail with reference to thefollowing Examples and Test Examples, which, however, are not intendedto restrict the scope of the invention.

Example 1 Production of bis(2-aminomethylpyridine)/zinc(II) complex[Zn(2-AM-py)₂(H₂O)Cl₂]

An ethanol solution of zinc(II) chloride (5 mmols) was dropwise added toan ethanol solution of 2-aminomethylpyridine (10 mmols) with stirring.After this was left overnight, the resulting precipitate was taken outthrough filtration, washed with ethanol and ether, and dried to obtain awhite product as entitled.

Yield: 30%. m.p.: 197-199° C.

Molecular formula: Zn(C₆H₈N₂)₂Cl₂.0.9H₂O,

Molecular weight: 368.8.

Elementary analysis:

cald. (%) C; 39.08, H; 4.86, N; 15.19 found (%) C; 39.07, H; 4.49, N;15.20,

IR (KBr): full chart in FIG. 4.

In the same manner as above, produced were bis-(R)- and(S)-2-(1-aminoethyl)pyridine (=α-pyridylethylamine/zinc(II) complex[Zn((R)-2-(1-AE)-py)₂(H₂O)Cl₂ and Zn((S)-2-(1-AE)-py)₂(H₂O)Cl₂].Bis-(R)- and (S)-2-(1-aminoethyl)pyridine was produced according to areference (Uenishi et al. Heterocycles, 52, 719 (2000)).

R-Form

Yield: 64%. m.p.: 163-165° C. [α]_(D): +1.2°.

Molecular formula: Zn(C₇H₁₀N₂)₂Cl₂.0.2H₂O,

Molecular weight: 384.2.

Elementary analysis:

cald. (%) C; 43.76, H; 5.34, N; 14.54 found (%) C; 43.92, H; 5.34, N;14.54.S-Form

Yield: 77%. m.p.: 162-164° C. [α]_(D): −1.2°.

Molecular formula: Zn(C₇H₁₀N₂)₂Cl₂.0.5H₂O,

Molecular weight: 389.6.

Elementary analysis:

cald. (%) C; 43.16, H; 5.43, N; 14.38 found (%) C; 43.22, H; 5.36, N;14.43.

Example 2 Production of N,N′-ethylene-bis-β-alanine/zinc(II) complex[Zn(βAeAβ)(H₂O)₂]

N,N′-ethylene-bis-β-alanine (10 mmols) was dissolved in an aqueoussolution of barium hydroxide (5 mmols). To the resulting solution,dropwise added was an aqueous solution of zinc(II) sulfate (5 mmols)with stirring. After left for 1 day, the resulting precipitate wasseparated through filtration, and the filtrate was concentrated. Theresulting white precipitate was recrystallized from hot water to obtaina white product as entitled.

Yield: 92%.

IR (KBr): ν C═O; 1568 cm⁻¹.

Molecular formula: Zn(C₈H₁₄N₂O₄)(H₂O)₂,

Molecular weight: 306.2.

Elementary analysis:

cald. (%) C; 29.58, H; 5.46, N; 9.86 found (%) C; 29.33, H; 5.39, N;9.76.

Example 3 Production of N,N′-trimethylene-bis-L-valine/zinc(II) complex[Zn(VtV)(H₂O)₂]

N,N′-ethylene-bis-L-valine (5 mmols) was dissolved in an aqueoussolution of barium hydroxide (10 mmols). To the resulting solution,dropwise added was an aqueous solution of zinc(II) sulfate (5 mmols)with stirring. After left for 1 day, the resulting precipitate wasseparated through filtration, and the filtrate was concentrated. Theresulting white precipitate was recrystallized from hot water to obtaina white product as entitled.

Yield: 79%.

IR (KBr): ν C═O; 1591 cm⁻¹. S-form: [α]_(D): +23.3°.

Molecular formula: Zn(C₁₃H₂₄N₂O₄)(H₂O)₂,

Molecular weight: 397.2.

Elementary analysis:

cald. (%) C; 39.31, H; 7.97, N; 7.05 found (%) C; 39.21, H; 7.53, N;7.03.

Example 4 Production of L-aspartyl-L-phenylalanine methyl ester(aspartame)/zinc(II) complex [Zn(AspF—OMe)(H₂O)₂]

Ba(OH)₂/8H₂O (5 mmols) was added to an aqueous solution of aspartame (10mmols). To the resulting solution, dropwise added was an aqueoussolution of zinc(II) sulfate (5 mmols) with stirring. After left for 1day, the resulting precipitate was separated through filtration, and thefiltrate was concentrated. This was dried in vacuum to obtain a whiteproduct as entitled.

Yield: 62%. S-form: [α]_(D): +11.6°.

IR (KBr): ν C═O (amido); 1655 cm⁻¹, ν C═O (ester); 1742 cm⁻¹.

Molecular formula: Zn(C₁₃H₂₄N₂O₄)(H₂O)₂,

Molecular weight: 695.2.

Elementary analysis:

cald. (%) C; 48.37, H; 5.63, N; 8.06 found (%) C; 48.38, H; 5.48, N;8.19.

Example 5 Production of bis(glycyl-N,N′-ethylene-L-alanyl-L-alanineethyl ester)/zinc(II) complex [Zn(Gly-eAA-OEt)₂Cl₂]

Barium hydroxide (5.5 mmols) dissolved in a small quantity of water wasadded to a water/methanol (1:1) solution withglycyl-N,N′-ethylene-L-alanyl-L-alanine ethyl ester monohydrochloride(11 mmols) dissolved therein. To the resulting solution, dropwise addedwas an aqueous solution of zinc(II) sulfate (5 mmols) with stirring.After left for 1 day, the resulting precipitate was separated throughfiltration, and the filtrate was concentrated. This was dried to obtaina white product as entitled.

Yield: 82%. m.p.: 102-110° C. [α]_(D): +59.8°.

IR (KBr): ν C═O (amido); 1638 cm⁻¹, ν C═O (ester); 1734 cm⁻¹.

Molecular formula: Zn(C₁₂H₂₁N₃O₄)₂Cl₂.4H₂O,

Molecular weight: 751.0.

Elementary analysis:

cald. (%) C; 38.38, H; 6.71, N; 11.19 found (%) C; 37.92, H; 6.64, N;11.56.

Glycyl-N,N′-ethylene-L-alanyl-L-alanine ethyl ester monohydrochloride(Gly-eAA-OEt/HCl) used herein was prepared by producing Boc-Gly-eAA-OEtaccording to a reference, T. Yamashita, Y. Kojima, K. Hirotsu, A.Ohsuka, Int. J. Peptide Protein Res., 33, 110 (1989), and deprotectingit with 4 N HCl/AcOEt.

Example 6 Production of N,N-dipyridylmethyl-L-valine/zinc(II) complex[Zn(^(pm2)V)(ClO₄)]

Barium hydroxide (5 mmols) dissolved in a small quantity of water wasadded to a water/methanol (1:1) solution withN,N-dipyridylmethyl-L-valine methyl ester monohydrochloride (10 mmols)dissolved therein. To the resulting solution, dropwise added was anaqueous solution of zinc(II) sulfate (10 mmols) with stirring. Afterthis was left for 1 hour, barium perchlorate (5 mmols) was added theretoand stirred overnight. The resulting precipitate was separated throughfiltration, and the filtrate was concentrated. This was dried to obtaina white product as entitled.

Yield: 87%. m.p.: 208-220° C. [α]_(D): −101.0°.

IR (KBr): ν C═O (amido); 1638 cm⁻¹, ν C═O (ester); 1734 cm⁻¹.

Molecular formula: Zn(C₁₇H₂₀N₃O₆)ClO₄.1.8H₂O,

Molecular weight: 524.5.

Elementary analysis:

cald. (%) C; 38.38, H; 6.71, N; 11.19 found (%) C; 37.92, H; 6.64, N;11.56.

In the same manner as above, produced were zinc(II) complexes withN,N-dipyridylmethyl-D-valine and N-6-methylpyridylmethyl-L-aspartic acid[Zn(^(pm2)V_(R))(ClO₄) and Zn(^(6Me-pm)D)].

N,N-dipyridylmethyl-D-valine/zinc(II) complex

Yield: 42%. m.p.: 210-218° C. [α]_(D): +107.4°.

IR (KBr): ν C═O (amido); 1638 cm⁻¹, ν C═O (ester); 1734 cm⁻¹.

Molecular formula: Zn(C₁₇H₂₀N₃O₆)Cl₂.1.8H₂O,

Molecular weight: 495.6.

Elementary analysis:

cald. (%) C; 41.20, H; 4.80, N; 8.48 found (%) C; 40.94, H; 4.21, N;8.38.

N-6-methylpyridylmethyl-L-aspartic acid/zinc(II) complex

Yield: 82%. m.p.: 279-283° C.

IR (KBr): ν C═O; 1590 cm⁻¹.

Molecular formula: Zn(C₁₁H₁₀N₂O₄).2.7H₂O,

Molecular weight: 348.2.

Elementary analysis:

cald. (%) C; 37.94, H; 4.46, N; 8.04 found (%) C; 38.02, H; 4.70, N;8.07.

Ester, hydrochloride with N,N-dipyridylmethyl-L-valine,N,N-dipyridylmethyl-D-valine and N-6-methylpyridylmethyl-L-aspartic acidhydrochloride used herein were prepared and saponified according to themethod described in a reference, K. Yamato et al., Chem. Lett., 1999,295.

Example 7 Production of L-lactic Acid (Lac)/zinc(II) complex [Zn(Lac)₂]

An aqueous solution of zinc sulfate (5 mmols) was dropwise added to anaqueous solution of lactic acid (10 mmols) and lithium hydroxide (10mmols). This was kept stirred overnight, and the resulting precipitatewas taken out through filtration and washed with water to obtain theintended product.

Yield: 90%. m.p.: >300° C. [α]_(D): −11.1° (CH₃OH).

Molecular formula: Zn(C₆H₃₀O₆).2H₂O,

Molecular weight: 279.6.

Elementary analysis:

cald. (%) C; 25.78, H; 5.05 found (%) C; 25.87, H; 5.05.

Example 8 Production of bis(quinic acid)/zinc(II) complex [Zn(Qui)₂]

An aqueous solution of zinc(II) sulfate (2.5 mmols) was added to anaqueous solution of quinic acid (5 mmols) and barium (2.5 mmols). Theresulting precipitate was separated through filtration, the filtrate wasconcentrated, and the resulting residue was reprecipitated fromwater/methanol to obtain the intended product.

Yield: 49%. m.p.: >300° C. [α]_(D): +207° (H₂O).

Molecular formula: Zn(C₁₄H₂₂O₁₂)/0.2H₂O,

Molecular weight: 451.3.

Elementary analysis:

cald. (%) C; 37.26, H; 5.00 found (%) C; 37.24, H; 4.84.

Example 9 Production of L-carnitine(γ-trimethyl-β-hydroxybutyrobetaine=Car)/zinc(II) complex [Zn(Car)₂Cl₂]

An aqueous solution of zinc(II) chloride (5 mmols) was dropwise added toan aqueous solution of L-carnitine (10 mmols) with stirring. Afterstirred for 2 hours, the reaction liquid was concentrated, and theresidue was processed with ethanol to obtain the intended product,powdery crystal.

Yield: 88%. m.p.: 173-175° C. [α]_(D): +3.2° (H₂O).

Molecular formula: Zn(C₁₄H₃₀N₂O₆)Cl₂/H₂O,

Molecular weight: 476.7.

Elementary analysis:

cald. (%) C; 35.27, H; 6.77, N; 5.88 found (%) C; 35.04, H; 6.82, N;5.67,

IR (KBr): full chart in FIG. 5.

Example 10 Production of bis(L-ascorbic acid)/zinc(II) complex[Zn(Vit-C)₂]

An aqueous solution of zinc(II) chloride (5 mmols) was added to anaqueous solution of L-ascorbic acid (10 mmols) and lithium hydroxide (10mmols) with stirring. After left overnight, this was filtered, and thefiltrate was concentrated. The insoluble substance was separated inethanol through filtration, and the resulting filtrate was concentrated.The residue was processed with ether to obtain the intended product,powdery crystal.

Yield: 55%. m.p.: 158-198° C. (decomposed). [α]_(D): −162.9° C. (H₂O).

Molecular formula: Zn(C₆H₇O₆).1.4H₂O,

Molecular weight: 440.9.

Elementary analysis:

cald. (%) C; 32.69, H; 3.84 found (%) C; 32.89, H; 4.12,

IR (KBr): full chart in FIG. 6.

Example 11 Production of bis(vitamin U)/zinc(II) complex [Zn(Vit-U)Cl₂]

A methanol solution of zinc(II) chloride (10 mmols) was dropwise addedto a methanol solution of vitamin U (10 mmols) and lithium hydroxide (10mmols) with stirring. After this was left overnight, the precipitateformed was taken out through filtration, and the resulting white powderycrystal was washed three times with methanol to obtain the intendedproduct.

Yield: 93%. m.p.: 117-120° C. [α]_(D): −0.25° (H₂O).

Molecular formula: Zn(C₆H₁₃NO₂S)Cl₂/0.75H₂O,

Molecular weight: 313.0.

Elementary analysis:

cald. (%) C; 23.02, H; 4.67, N; 4.47 found (%) C; 22.93, H; 4.40, N;4.54,

IR (KBr): full chart in FIG. 7.

Example 12 Production of bis(L-theanine)/zinc(II) complex [Zn(Tea)₂]

A methanol solution of zinc(II) nitrate.6H₂O (5 mmols) was dropwiseadded to a methanol solution of L-theanine (L-glutamine-monoethylamide)(10 mmols) and lithium hydroxide (10 mmols) with stirring. After thiswas left for 2 hours, the precipitate formed was taken out throughfiltration, and the resulting white powdery crystal was washed threetimes with methanol.

Yield: 99%. m.p.: >300° C. [α]_(D): −6.4° (H₂O).

Molecular formula: Zn(C₁₄H₂₆N₄O₆),

Molecular weight: 411.8.

Elementary analysis:

cald. (%) C; 40.84, H; 6.36, N; 13.61 found (%) C; 40.89, H; 6.33, N;13.46,

IR (KBr): full chart in FIG. 8.

Pharmaceutical Test Example 1

According to the method described in Biol. Pharm. Bull., 18, 719-725(1995), the following experiment was carried out.

Isolation of Rat Fat Cells:

While anesthetized with ether, male Wistar rats (body weight, 200 g)were bled to death, and the fat cells were isolated from the fat tissuearound the epididymis thereof, according to Rodbell's method (J. Biol.Chem., 239, 375 (1964)). The fat cells were cut with scissors, anddigested in a KRB buffer (glucose 10 mM, NaCl 120 mM, CaCl₂ 1.27 mM,MgSO₄ 1.2 mM, KCl 4.75 mM, KH₂PO₄ 1.2 mM and NaHCO 24 mM, pH=7.4)containing 20 mg/ml of bovine serum albumin (BSA) and 2 mg/ml ofcollagenase, at 37° C. for 1 hour. Filtered through a nylon mesh (250mm), the fat cells were separated from the non-digested tissue, and thenwashed three times with the buffer mentioned above but not containingthe collagenase to prepare a cell suspension of 2.5×10⁶ cells/ml.

Effect of Zinc(II) Complex on Rat Fat Cells:

In silicon-processed vials, the fat cells isolated in the above (2.5×10⁶cells/ml) were pre-incubated in 1 ml of KRB buffer that contains avarying concentration (10⁻⁴, 5×10⁻⁴, 10⁻³ M) of VOSO₄ different Zn(II)complexes of the invention and 20 mg/ml of BSA, at 37° C. for 0.5 hours.Next, 10⁻⁵ M of epinephrine was added to the reaction mixture, and theresulting solution was incubated at 37° C. for 3 hours. The reaction wasstopped by cooling the mixture with ice, and the mixture was centrifugedat 3000 rpm for 10 minutes. The free fatty acid (FFA) level in theextracellular solution was measured with an NEFA kit, and IC₅₀ of thetest compound was calculated.

The results are given in FIG. 1, FIG. 2, FIG. 3 and Table 1 (mentionedabove).

Pharmaceutical Test Example 2

<Test Method>

Type II diabetic model animals, KK-A^(y) mice (8 weeks old), were usedin the test. To those of a control group, administered was a 5% acaciasolution (n=6). Any of Zn(2-AM-py)₂Cl₂, Zn(Lac)₂, Zn(Qui)₂, Zn(Car)₂Cl₂and a mixed solution of zinc sulfate and vitamin U was dissolved in 5%acacia solution to prepare complex solutions. To those of a test group,administered was the complex solution (n=6). The dose was 2 mg Zn/kg to3 mg Zn/kg. The acacia solution alone or the complex solution wasintraperitoneally administered to each mouse once a day (but Zn(Car)₂Cl₂was orally administered). The dose of the mixed solution of zinc sulfateand vitamin U was 3 mg Zn/kg for 3 days after the start of theadministration, and on day 4 and thereafter, the dose was varieddepending on the blood glucose level in each mouse (5 mg Zn/Kg to thosehaving a blood glucose level of 200 mg/dl or more; and 3 mg Zn/kg tothose having a blood glucose level of less than 200 mg/dl). Thisexperiment was intraperitoneally administered to each mouse once a day.

To the mice of the control group, administered was 0.5 ml of the acaciasolution alone.

The presence or absence of diabetes in the tested mice was confirmed bythe average blood glucose level of at least 450 mg/dl and the averagebody weight of at least 35 g before administration.

The blood glucose level was measured with a simple blood glucose levelmeter (Glucocard, by Arclay Factory, Kyoto).

<Result>

FIG. 9 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of Zn(2-AM-py)₂Cl₂ for 14 days (-o-) andwith no administration thereof (control) (-•-).

FIG. 10 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of Zn(Lac)₂ for 14 days (-o-) and with noadministration thereof (control) (-•-).

FIG. 11 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of Zn(Qui)₂ for 14 days (□) and with noadministration thereof (control) (▪).

FIG. 12 shows the blood glucose level (BGL) change in KK-A^(y) mice withoral administration of Zn(Car)₂Cl₂ for 14 days (-o-) and in those withoral administration of L-carnitine for 14 days (-♦-).

FIG. 13 shows the blood glucose level (BGL) change in KK-A^(y) mice withintraperitoneal administration of a solution (about pH 7) prepared bymixing zinc sulfate and vitamin U in a molar ratio of 1:2 (that is, asolution of Zn(Vit-U)Cl₂)) for 14 days (-o-) and with no administrationthereof (control) (-•-).

FIG. 14 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of Zn(2-AM-py)₂Cl₂ for 14 days (-o-) andwith no administration thereof (control) (-•-).

FIG. 15 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of Zn(Lac)₂ for 14 days (-o-) and with noadministration thereof (control) (-•-).

FIG. 16 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of Zn(Qui)₂ for 14 days (-o-) and with noadministration thereof (control) (-•-).

FIG. 17 shows the body weight change of KK-A^(y) mice with oraladministration of Zn(Car)₂Cl₂ for 14 days (-o-) and of those with oraladministration of L-carnitine for 14 days (-♦-).

FIG. 18 shows the body weight change of KK-A^(y) mice withintraperitoneal administration of a solution (about pH 7) prepared bymixing zinc sulfate and vitamin U in a molar ratio of 1:2 (that is, asolution of Zn(Vit-U)Cl₂)) for 14 days (-o-) and with no administrationthereof (control) (-•-).

<Conclusion>

As is obvious from FIG. 9 to FIG. 13, the mice of the test groups, towhich any of five different types of zinc(II) complexes of the inventionhad been administered, all enjoyed blood glucose level normalization ascompared with those of the control groups.

Also obvious from FIG. 14 to FIG. 18, body weight reduction, one indexof side effects was not almost seen in the mice of the test groups towhich the zinc(II) complex of the invention had been administered.

Pharmaceutical Test Example 3

This is a glucose load test effected according to the method describedin Biochem. Biophys. Res. Comm., 281, 1190-1193 (2001), in which thezinc(II) complex of the invention was intraperitoneally administered toKK-A^(y) mice for 14 days or was not thereto. The mice were fed withnothing for 13 days, then 1 g/kg of glucose was orally administered tothem, and the blood glucose level in them was measured at regularintervals.

<Result>

FIG. 19 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with intraperitoneal administration ofZn(2-AM-py)₂Cl₂ for 14 days (-o-) and with no administration thereof(control) (-•-). The glucose tolerance test was effected after the 14days administration.

FIG. 20 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with oral administration of Zn(Car)₂Cl₂ for 14days (-o-), with oral administration of L-carnitine for 14 days (-♦-),and with administration of acacia solution alone for 14 days (control)(-•-). The glucose tolerance test was effected after the 14 daysadministration.

FIG. 21 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with intraperitoneal administration of Zn(Qui)₂for 14 days (-o-) and with no administration thereof (control) (-•-).The glucose tolerance test was effected after the 14 daysadministration.

FIG. 22 shows the blood glucose curve observed in a glucose tolerancetest in KK-A^(y) mice with intraperitoneal administration of a solution(about pH 7) prepared by mixing zinc sulfate and vitamin U in a molarratio of 1:2 (that is, a solution of Zn(Vit-U)Cl₂)) for 14 days (-o-)and with no administration thereof (control) (-•-). The glucosetolerance test was effected after the 14 days administration.

<Conclusion>

As is obvious from FIG. 19, the mice with 13-hour fasting after 14-dayintraperitoneal administration of Zn(2-AM-py)₂Cl₂ had a low peak top ofblood glucose level after glucose administration thereto, as comparedwith those with no complex administration. In the former, the bloodglucose level smoothly lowered and after 120 minutes, it reached almostthe normal level.

Also obvious from FIG. 20, the mice with 13-hour fasting after 14-dayintraperitoneal administration of Zn(2-AM-py)₂Cl₂ had a low peak top ofblood glucose level after glucose administration thereto, as comparedwith those with 13-hour fasting after 14-day oral administration ofL-carnitine and those with 14-day administration of acacia solutionalone. In the former, the blood glucose level smoothly lowered and after120 minutes, it reached almost the normal level.

Further obvious from FIG. 21 and FIG. 22, the mice with 14-dayintraperitoneal administration of Zn(Qui)₂ or Zn(Vit-U)Cl₂ followed bythe glucose load test were significantly ameliorated in point of theirdiabetic condition than the diabetic mice (control mice), though theireffect was somewhat lower than that of the mice administered withZn(2-AM-py)₂Cl₂ or Zn(Car)₂Cl₂.

Example 13

30 g of water was mixed with 100 g of butter, 30 g of sugar, 200 g ofwheat, 0.5 to 5 g of sodium bicarbonate and 3 g of Zn(Lac)₂. Theresulting mixture was heated and baked into soft cookies.

Examples 14 to 18

Soft cookies were produced in the same manner as in Example 13, forwhich, however, 3 g of Zn(Qui)₂, Zn(Car)₂Cl₂, Zn(Vit-C)₂, Zn(Vit-U)Cl₂or Zn(Tea)₂ was used in place of 3 g of Zn(Lac)₂ in Example 13.

Example 19

Soft cookies were produced in the same manner as in Example 13, forwhich, however, 1 to 5 g of Zn(Lac)₂ and 1 to 20 g of DHA-containingfish oil were used in place of 3 g of Zn(Lac)₂ in Example 13.

Example 20

Various juices were produced by mixing 1 to 50 g of Zn(Vit-C)₂, pHcontrolling agent (sodium hydroxide), sweetener and powdery juice stock.

INDUSTRIAL APPLICABILITY

The zinc(II) complex of the invention is highly stable and hasfat-soluble insulin-like activity and hypotensive activity. Accordingly,the zinc(II) complex of the invention is extremely useful for medicinesthat are used for preventives/remedies for glucose tolerance disorders,diabetes (e.g., type II diabetes), insulin-resistant syndromes (e.g.,insulin receptor disorders), polycystic ovary syndromes, hyperlipemia,atherosclerosis, cardiovascular disorders (e.g., stenocardia, cardiacinsufficiency), hyperglycemia, hypertension, stenocardia, pulmonaryhypertension, congestive cardiac insufficiency, diabetic complications(e.g., diabetic gangrene, diabetic arthropathy, diabeticglomerulosclerosis, diabetic skin disorders, diabetic neuropathy,diabetic cataract, diabetic retinopathy), taste disorders, skindisorders, etc. In addition, it is much expected for health(supplementary) foods and nutrient (supplementary) foods that areeffective for prevention and remedy of insulin and blood glucose-relateddisorders.

1. A method for lowering blood glucose level in a subject comprisingadministering to the subject a zinc(II) organic complex having, as aligand, a compound of the formula (1) (when they are optically activecompounds, they have both (R)-form and (S)-form thereof):

wherein R, R′, R¹ and R² each independently represents a hydrogen atom,an alkyl group, an aryl group, or an aralkyl group; and n indicates aninteger of from 1 to
 3. 2. The method according to claim 1, wherein thezinc(II) organic complex is administered orally.